Model organisms, such as Saccharomyces cerevisiae, have proven effective in elucidating genes and metabolites that modulate life span and this knowledge forms a basis for understanding human aging and longevity. These studies have identified the Akt/PKB protein kinases as conserved regulators of aging. However, it is not entirely clear how they work. One focus of our research is the S. cerevisiae Sch9 protein kinase, an AKT homolog that regulates chronological life span (CLS). From in vitro studies we identified sphingolipid long chain bases (LCBs) and Pkh1, a homolog of human phosphoinositide-dependent protein kinase 1 (PDK1), as the first upstream regulator of Sch9. In addition, we identified the first Sch9 substrates, Rli1, involved in translation initiation and ribosome biogenesis, and Ahp1, an alkyl hydroperoxide reductase that protects against oxidative stress. We will build upon these novel results and determine if phosphorylation of Rli1 and Ahp1 by Sch9 plays a role in CLS. We will also determine in vivo if LCBs and Pkh1 act upstream to regulate Sch9 during chronological aging. Although we have successfully identified Sch9 substrates, there are likely to be others and we propose to search for new substrates by classical biochemical strategies, newer automated but complementary strategies, and a genetic screening strategy. In summary, the results of these studies will provide the much needed mechanistic understanding of how Akt (Sch9) activity is regulated in yeast and how it regulates CLS through downstream substrates. This knowledge will help to provide a molecular basis for understanding human aging and longevity. PUBLIC HEALTH RELEVANCE: The proposed research will determine how the Sch9 protein kinase, a functional homolog of mammalian Akts/PKBs, regulates chronological life span in the Baker's yeast Saccharomyces cerevisiae. Signal transduction pathways that contain Akt/PKB type protein kinases play important roles in regulating aging and life span, and these pathways have been conserved from yeast to mammals. Thus, information gained from these studies should provide a more mechanistic basis for understanding how aging and life span are regulated in humans.